Expression And Identification Of Recombinant Adenovirus Mediated Macaca Mulatta CD40L Extracellular Period Of Gene In Macaca Mulatta Immature Dendritic Cells

[Object]Expression and identification of recombinant adenovirus mediated macaca mulatta CD40L extracellular period of gene in macaca mulatta immature dendritic cells.[Methods]1Application of HEK293cell packing pAd-CMV-sCD40L adenovirus vector, and deno-x purification kit purified virus。2)Utilize lymphocyte separation medium separation of macaca mulatta mononuclear cells;3) Apply magnetic bead method separation CD34+positive cells in macaca mulatta mononuclear cells, These cells can be induced into immature dendritic cells by the application of IL-4and GM-CSF, and through identify the purity and maturity by flow cytometry;4) Utilize recombinant adenovirus vector infect of macaca mulatta immature dendritic cells;5) PCR and Western Blot appraisal CD40L expression in immature dendritic cells;6) Mixed lymphocyte culture. The proliferation capacity of allogeneic T cells stimulated by different treatment groups of dendritic cells was examined by CCK-8Kits;[Result]PT-PCR amplification, PCR products by electrophoresis, displays the band of170bp, matched with the length of target gene theory band. Western bloting display23KD of band matched with CD40L protein size. In MLR (same-reaction radio), the capacity of the group of imDCs were infected recombinant adenovirus for stimulating the proliferation of T-cells was remarkably lower than the other groups.[Conclution]macaca mulatta CD40L extracellular period of gene can be stable expression in immature dendritic cells. The CD40L protein can block the CD40-CD40L pathway, thereby inhibiting the proliferation of T cells. [Prospect]In the macaca mulatta liver transplantation, the imDCs, witch infected by recombinant adenovirus, can be injected in receptor macaca mulatta’s portal vein. It is expected to be effective in inducing macaca mulatta liver transplantation immune tolerance.